The mechanism by which a protein folds to its native, biologically active structure is one of the major unsolved problems of molecular bioscience, with important practical consequences for rational protein design, protein structure prediction and folding related disease states. The long term objective of this proposal is to characterize the molecular dynamics and mechanisms of early events in protein folding. Fundamentally new approaches to the rapid initiation and characterization of folding reactions are required to fulfill this objective. The viability of such an approach has been established in experiments using a laser-induced temperature jump and time-resolved infrared spectroscopy. Thus, for the first time, it is possible to initiate and follow the kinetics of protein folding or unfolding on a time scale of 50 ps, or some 10(7) times faster than conventional (rapid mixing) kinetic studies. The organizing question of this work is what are the dynamics and mechanisms of secondary structure formation in an isolated peptide and in a protein, and what is the role of secondary structure formation in guiding the folding process and stabilizing early intermediates. Specifically, new methods for the rapid initiation of protein folding will be developed, using a laser-induced, impulsive macroscopic perturbation of temperature or pH. Time-resolved infrared spectroscopy will be used as a structure specific probe of the dynamics of protein folding intermediates. The amide vibrations of peptides and proteins, together with isotopic labeling will be used to identify specific secondary structures in static and time-resolved infrared spectra. A hierarchy of problems will be investigated. Starting with a series of de novo helical peptides, the dynamics and mechanism of helix formation will be investigated. The rate of helix nucleation and propagation, and the role of short range interactions (electrostatic, salt bridges, end capping) will be explored. Second, the dynamics and mechanism of folding of fragments of the fast folding core of myoglobin will be explored. The influence of short range interactions is again the focus, particularly the formation of reverse turns and helices, and their relative roles in the folding reactions of small peptides. Finally, the whole protein (apomyoglobin) will be investigated once the dynamics and infrared signatures of secondary structural elements are established, to probe the relative role of secondary structure formation versus long range interactions (hydrophobic collapse) in controlling the course of a folding reaction.